Since BC is edible as well as tasteless and odorless, it is utilized in the food industry. BC's high dispensability in water further provides it with a lot of industrial applications, such as to maintain moisture and viscosity of food, cosmetics or coating agents, to strengthen food materials, to improve stability of food, and to be used as low-calorie additives and an emulsion stabilizer.
BC is characterized by a sectional width of its fibrils which is smaller by two orders of magnitude than that of other kinds of cellulose such as those derived from wood pulp.
Owing to such structural and physical feature of microfibril, a homogenized BC has plenty of industrial applications as a strengthening agent for polymers, especially hydrophilic polymers. Products prepared by solidification of the macerated BC in the form of a lump or paper show a high elastic modulus in tension owing to the above feature, and are therefore expected to have excellent mechanical properties for use in various kinds of industrial materials.
The strains conventionally used in the production of BC include Acetobacter strains such as Acetobacter xylinum subsp. sucrofermentans such as the BPR 2001 strain, Acetobacter xylinum ATCC23768, Acetobacter xylinum ATCC23769, Acetobacter pasteurianus ATCC10245, Acetobacter xylinum ATCC14851, Acetobacter xylinum ATCC11142, Acetobacter xylinum ATCC10821; and strains derived and bred from those strains by means of various kinds of mutagenesis treatment and recombination of genes; and strains derived and generated from those strains by using known mutagens such as NTG (nitrosoguanidine).
The taxonomic characters of BPR 2001 strain are as follows:
Morphology: rod, Gram stain: negative, Spore formability: negative, Behavior toward oxygen: aerobic, Catalase: positive, Oxidase: negative, Formation of acetic acid from ethanol: positive, Oxidation of acetates: positive, Oxidation of lactates: positive.
The PQQ non-generating strain obtained from the BPR 2001 strain has been also used. One example of the above PQQ non-generating strain, designated the BPR 3001c was deposited at the National Institute of Bioscience and Human-Technology, Agency of Industrial Science and Technology (1-3, Higashi 1-chome, Tsukuba-shi, Ibaraki-ken 305 Japan) on May 2, 1994 under accession number FERM P-14297, and then transferred on May 12, 1995 to the deposit under the terms of the Budapest Treaty on the International Recognition of the Deposit of Microorganisms for the Purposes of Patent Procedure and Regulation under accession number FERM BP-5100.
Mutants include a levan sucrase-defective mutant in which the production of levan is suppressed.
Other mutants include the sulfur agent-resistant strain designated BPR 3001D, pyrimidine analogue-resistant strain designated BPR 3001I and DHO-DHase inhibitors-resistant strain designated BPR3001N, which were also deposited at the National Institute of Bioscience and Human-Technology, Agency of Industrial Science and Technology on May 25, 1994 under accession number FERM P-14330, on Jun. 10, 1994 under accession number FERM P-14362, and on Jun. 10, 1994 under accession number FERM P-14361, respectively. Further, there have been disclosed the cellulose-producing bacteria transformed with a gene for an enzyme involved in sucrose metabolism (WO95/32279) and the cellulose-producing bacteria transformed with an extracellular invertase gene and its secretion-accelerating gene (Japanese Patent Application Hei 7 (1995)-252021).
BPR 2001 was deposited at the National Institute of Bioscience and Human-Technology, Agency of Industrial Science and Technology (1-3, Higashi 1-chome, Tsukuba-shi, Ibaraki-ken 305 Japan) on Feb. 24, 1993 under accession number FERM P-13466, and then transferred on Feb. 7, 1994 to the deposit under the terms of the Budapest Treaty on the International Recognition of the Deposit of Microorganisms for the Purposes of Patent Procedure and Regulation under accession number FERM BP-4545.
The present inventors have carried out many kinds of researches in order to provide novel cellulose-producig bacteria for the production of a large amount of BC using sucrose or glucose as sugar source, in an efficient manner, and have found a novel subspieces which is substantialy negative or very slightly positive in oxidation of acetates and lactates.
The cellulose-producing bacteria grow and produce BC, while incorporating and metabolizing carbon sources such as saccharides, and biosynthesizing amino acids or incorporating and metabolizing them form a culture medium. The present inventors have therefore carried out many kinds of researches, taking into consideration the increase of capability of incorporation and metabolism of saccharides and amino acids, so that strains which are improved in the cellulose production may be obtained. As a result, the present inventors have surprisingly found that the additon of some compounds (saccharides analogs or amino acid analogs) could prevent the growth of the bacteria. The present inventors have obtained strains with an improved productivity of BC by selecting the resistant strains against these analogs, and completed the present invention.
Levan sucrase (EC2.4.1.10) is known to decompose and metabolize sucrose. This enzyme has two activities, (1) hydrolysis activity of sucrose into glucose and fructose, and (2) transfructosylation activity to produce glucose and levan from sucrose. The latter activity is not preferred in terms of BC production since it will produce levan as a by-product. It will be very advantageous in the production and purification of BC, if the accumulation of levan is suppressed. The present inventors have already disclosed in the Japanese Patent Application Hei 7 (1995)-252021 that by deriving the levan sucrase-defective strain and incorporating the gene of an extracellular invertase or a levan sucrase with a reduced levan productibity into said strain, the accumulation of levan will be reduced and the cellulose may be efficiently produced.